Abstract

Immune responses primed by endogenous heat shock proteins (HSPs), specifically gp96, have been used for immunotherapy of cancer. Immunization with low doses of gp96 primes a response that is dominated by T helper type 1 (Th1) cells and cytotoxic T lymphocytes (CTLs), which allows for recognition and killing of tumor cells. However, increasing the dose of gp96 primes a response characterized by regulatory T (Treg) cells and immunosuppression. Although the T cell responses driving this dose dichotomy have been studied in multiple systems, innate mechanisms that control T cells in the context of gp96 immunization remains unknown. The antigen presenting cells (APCs) involved in this response are the main focus of Chapters 1 and 2 of this thesis. Here we show gp96 preferentially engages conventional and plasmacytoid dendritic cells (pDCs) under low and high doses respectively, through the HSP receptor CD91. We show that DNA methyltransferase 1 (DNMT1) is active in multiple APC population in response to gp96 stimulation, and that global DNA methylome and protein expression changes occur in these populations in response to in vivo immunization. Methylation-dependent upregulation of neuropilin-1 (Nrp1) on pDCs enables long term interactions with Treg cells that enhance their function in suppressing ongoing Th1 anti-tumor immunity. Our study defines a CD91-dependent mechanism through which gp96 controls dichotomous immune responses, relevant to the therapy of cancer and autoimmunity.Additionally, we are interested in the signaling pathways initiated by gp96-CD91 interaction. Previous studies have shown that NF-B and p38 MAPK are activated in HSP-responsive macrophages, but the consequences of p38 activation were not investigated. Chapter 3 will focus on the downstream mediators of p38, specifically the transcription factor STAT1, and the cytokine expression and secretion that follows. We show that gp96-treated macrophages upregulate STAT1-dependent cytokines as a result of STAT1 phosphorylation. This event requires intact CD91 and p38 MAPK. Importantly, STAT1 target CXCL10 is upregulated in response to gp96 and is critical for APC-NK cell crosstalk. Given that NK cells are required for anti-tumor immunity elicited by gp96 immunization, we hypothesize that STAT1 activation in responding APCs is necessary for tumor rejection.